|Budget Amount *help
¥3,100,000 (Direct Cost: ¥3,100,000)
Fiscal Year 1998: ¥1,600,000 (Direct Cost: ¥1,600,000)
Fiscal Year 1997: ¥1,500,000 (Direct Cost: ¥1,500,000)
To elucidate the physiological role of membrane-bound aminopeptidase of the larval midgut of the silkworm, Bombyx mori, especially in digestion and absorption of protein in an artificial diet, following three points were investigated. 1) longitudinal localization of enzyme in the midgut tissue, 2) Establishment of method of purification of the enzyme 3) Effect of protein content in an artificial diet on the enzyme activity.
The results were as follows.
1. Midgut tissue can be divided into three portions, namely, anterior, middle and posterior ones. Among them, the posterior portion was highest in activity per individual tissue basis, followed by the rest two portions with almost equal activities. This suggests that posterior portion plays a significant role in digestion and absorption of protein in the midgut.
2. A purification method was established. It employs solubilization of enzyme from midgut membrane fraction using an enzyme, phosphatidylinositol specific phospholipase C, followed by column chromatographies using separationmedia of ion exchange, metal chelate, and gel filtration.
3. The enzyme activity in the midgut of the silkworm larvae fed artificial diet containing protein of 6.4% and 13.2% in wet weight increased significantly compared to controls. This suggests that the enzyme is inducible upon feeding diet : with high content of protein.
The present study investigated an enzyme which is supposedly involved in digestion and absorption of protein in this insect. Another completely different approach is possible and we have a plan to do it shortly. It is a study of products analysis of protein digestion in the midgut tissue, absorbed peptides. if any, in the midgut cells, and absorbed peptides, if any, in the haemolymph in this insect. By identification of peptides of digestion products, we speculate the aproteases and peptidases that were involved in digestion. Then we search for the enzymes.